雷公藤内酯醇对浆细胞样树突状细胞功能及成熟的影响

目的 通过体外试验研究雷公藤内酯醇(triptolide)对浆细胞样树突状细胞(plasmacytoid dendritic cell,pDC)功能及成熟的影响,为进一步阐明雷公藤内酯醇的免疫学活性提供依据.方法 从健康志愿者外周血分离单个核细胞,流式细胞仪分选pDC,加入0、5、10、30 μg/L的雷公藤内酯醇共孵育,24 h后收集上清液,ELISA检测pDC分泌的IFN-α、IL-6、TNF-α量,5 d后收集细胞,流式细胞仪检测树突状细胞表型CD11c、CD80、CD86阳性率,光镜观察DC的形态,扫描电镜观察DC的超微结构.结果 雷公藤内酯醇显著降低pDC分泌的IFN-α、IL-6、TNF-α,并呈雷公藤内酯醇浓度依赖性(P＜0.05);雷公藤内酯醇可抑制pDC向DC的分化和成熟,并呈雷公藤内酯醇浓度依赖性(P＜0.05).结论 雷公藤内酯醇能够降低pDC的功能,并抑制其向DC的分化和成熟.

Abstract：

Objective To explore the mechanism of immunomodulatory activity of triptolide on healthy volunteers peripheral blood mononuclear cells (PBMC)-derived plasmacytoid dendritic cells (pDCs). Methods Healthy volunteers-derived pDCs were sorted by flow cytometry, then incubated with triptolide (0, 5, 10, 30 μg/L). After 24 hours, we detected the concentration of IFN-α, IL-6, TNF-α using ELISA. After 5 days, the cultrural cells were collected and analyzed by flow cytometry, light microscope and electron microscope scanning. Results Triptolide-treated pDCs secreted lower level of IFN-α,IL-6 ,TNF-α, triptolide could inhibit pDCs differentiation to DCs which displayed more immature morphology and immunophenotypes than untreated-pDCs. Conclusion Triptolide could decrease the immune function of pDCs, inhibit differentiation and maturation of pDCs.     

Engagement of BDCA-2 blocks TRAIL-mediated cytotoxic activity of plasmacytoid dendritic cells

Plasmacytoid dendritic cells (PDC) are a functionally distinct lineage of dendritic cells characterized by the release of large amounts of type I interferon (IFN I). IFN I release is efficiently triggered by viral infection and modulates several aspects of immune reactions including the activation of cytotoxic mechanisms finalized to the elimination of infected cells. In this study, we report that TLR7 and TLR9 ligands can induce the secretion of biologically active TNF-related apoptosis-inducing ligand (TRAIL) by PDC. Accordingly, PDC supernatant is endowed with TRAIL-mediated cytotoxic activity when tested on a TRAIL-sensitive Jurkat cell line. TRAIL production is only partially dependent on the autocrine production of IFN I as documented by the use of a blocking anti-IFNRA antibody and the stimulation with exogenous IFN I. Importantly, both TRAIL secretion and cytotoxic activity of PDC supernatants are completely abolished by BDCA2 ligation. These results provide further insights into the biological role of BDCA-2 and document a negative regulatory pathway of PDC cytotoxic activity that may be relevant in pathological situations such as tumors and autoimmune diseases.     

Histone deacetylase inhibitor valproic acid affects plasmacytoid dendritic cells phenotype and function

Objective

Plasmacytoid dendritic cells (PDC) represent a rare subset of dendritic cells specialized in the production of type I IFN in response to microbial pathogens. Recent data suggested that histone deacetylase (HDAC) inhibitors possess potent immunomodulatory properties both in vitro and in vivo. In this study, we assayed the ability of the HDAC inhibitor, valproic acid (VPA), to influence the phenotype and functional properties of human PDC isolated from peripheral blood.

Methods and results

We showed that VPA inhibited the production of IFN-α and the proinflammatory cytokines TNF-α and IL-6 by CpG-activated PDC. VPA also affected the phenotype of PDC by reducing the expression of costimulatory molecules induced by CpG activation. Moreover, VPA reduced the capacity of CpG-stimulated PDC to promote CD4⁺ T cell proliferation and IFN-γ production, while enhancing the proportion of IL-10 positive T cells.

Conclusion

These results suggest that HDAC inhibition by VPA alters essential human PDC functions, highlighting the need for monitoring immune functions in cancer patients receiving HDAC inhibitors, but also making these drugs attractive therapies in inflammatory, and autoimmune diseases implicating PDC.     

Severe herpes virus (HSV-2) infection in two patients with myelodysplasia and undetectable NK cells and plasmacytoid dendritic cells in the blood

BACKGROUND: How the immune system contains herpes simplex virus (HSV) infections is partly understood. T cells from infected persons proliferate in response to HSV antigens in vitro and may control local relapse rather than primary infection. NK cells have been involved in the control of experimental infections. A potentially important, as yet unexplored, population of interest might be the plasmacytoid dendritic cells (PDC), which contrary to monocytes, produce very high amounts of the major antiviral molecules, type-I interferon (IFN) following interaction with HSV. OBJECTIVES: Measure type-I IFN production, PDC, and NK cells in patients with unusually severe HSV infections. STUDY DESIGN: Two female patients of 33- and 50-year-old, respectively were referred because of severe disseminated HSV2 infection and myelodysplastic marrow. One patient had leukaemia and a primary HSV2 infection whereas the other had systemic lupus erythematosus (SLE) and a chronic HSV2 infection. The following studies were performed at various time points over 18 months: analysis of the lymphocytes and PDC subsets phenotype, lymphocyte proliferation assays to recall antigens; generation of NK cells in cultures, and production of type-I IFN in serum and by HSV-infected and by sendai virus (SV)-infected blood cells. RESULTS AND CONCLUSIONS: PDC and NK cells were undetectable in the blood of both patients and NK cells could not be generated in culture at the time of ongoing infection. PBMC failed to produce IFN after infection with HSV contrasting with a normal T cell proliferation to HSV antigens in patient 1. Our observation suggests that innate immunity, through NK cells and PDC may control HSV infections, and together with IFN-producing capacity, should be investigated in patients with unusually severe HSV infections.     

Requirement of HMGB1 and RAGE for the maturation of human plasmacytoid dendritic cells

Dendritic cells (DC) are key components of innate and adaptive immune responses. Plasmacytoid DC (PDC) are a specialized DC subset that produce high amounts of type I interferons in response to microbes. High mobility group box 1 protein (HMGB1) is an abundant nuclear protein, which acts as a potent pro-inflammatory factor when released extracellularly. We show that HMGB1 leaves the nucleus of maturing PDC following TLR9 activation, and that PDC express on the plasma membrane the best-characterized receptor for HMGB1, RAGE. Maturation and type I IFN secretion of PDC is hindered when the HMGB1/RAGE pathway is disrupted. These results reveal HMGB1 and RAGE as the first known autocrine loop modulating the maturation of PDC, and suggest that antagonists of HMGB1/RAGE might have therapeutic potential for the treatment of systemic human diseases.     

慢性HCV感染者外周血中髓样和浆样树突状细胞频数和表型研究

目的 探讨慢性HCV感染者外周血中髓样树突状细胞(mDC)和浆样树突状细胞(pDC)频数和表型的变化,并分析其与丙型肝炎临床指标间的相关性.方法 采用流式细胞术检测HCV感染者及健康对照外周血中mDC和pDC的频数及细胞表面共刺激分子HLA-DR、CD83、CD86、CD40和共抑制分子PD-L1的表达水平,并分析DC频数与HCV感染者血浆病毒载量、谷丙转氨酶(ALT)的相关性.结果 与健康对照组相比,HCV感染者外周血中mDC和pDC的频数明显降低(患者组分别为0.37±0.19和0.19±0.12,对照组为0.51±0.18和0.29±0.13,P＜0.05),且mDC频数与血浆HCV载量和血清ALT水平呈负相关(r=-0.5878,P＜0.0001;r=-0.4628,P=0.003).患者mDC和pDC表面共刺激分子HLA-DR、CD83、CD86、CD40以及共抑制分子PD-L1的表达均有不同程度升高,差别有统计学意义(共刺激分子P＜0.01,共抑制分子P＜0.05或0.01).结论 慢性HCV感染者外周血mDC和pDC频数下降,但DC表面共刺激分子和共抑制分子的表达均明显升高.该结果提示mDC数量的减少可能与HCV的慢性持续性感染有关.

Abstract：

Objective To explore the frequencies and phenotype of myeloid and plasmacytoid dendritic cells (mDC and pDC) in chronic HCV infection and to investigate the relationships between DC frequencies and HCV viral load and serum ALT level. Methods PBMC were isolated from chronic HCV infected patients and healthy control. Multi-color flow cytometry was used to analyze the frequencies and surface marker expression on mDC and pDC. The relationship between DC frequencies and viral load and ALT level was also calculated. Results In comparison with healthy control, frequencies of mDC and pDC in chronic HCV infection were significantly decreased (0. 37 ± 0. 19 and 0. 19 ± 0. 12 vs 0. 51 ± 0. 18 and 0. 29 ± 0.13, P＜0.05). The frequency of mDC was negatively correlated with HCV viral load (r= -0.5878, P ＜ 0. 0001 ) and serum ALT level ( r = - 0. 4628 , P = 0. 003 ). Both costimulatory markers ( HLA-DR, CD83, CD86, and CD40) and coinhibitory marker (PD-L1) expression on mDC and pDC in HCV infection were increased (P＜0.01 for costimulatory marker, P＜0.05 or F＜0.01 for coinhibitory marker). Conclusion The frequencies of mDC and pDC in chronic HCV infection were decreased, while the expression of costimulatory markers and coinhibitory marker were increased or not decreased in HCV infection. The decreased frequency of mDC was probably related to persistance of HCV infection.     

Functional dichotomy of plasmacytoid dendritic cells: antigen-specific activation of T cells versus production of type I interferon

Human plasmacytoid dendritic cells (PDC) are believed to link innate and adaptive immunity by producing type I interferon (IFN-I) and triggering adaptive T cell-mediated immunity. However, it remains elusive to which degree both PDC functions are linked. Here we show that CMV antigen targeted to PDC using a CD303 (blood dendritic cell antigen 2, BDCA-2) mAb is rapidly endocytosed and traffics via early sorting endosomes to emerging MHC-enriched compartments. Both processes occur independently of TLR ligand stimulation. Restimulation of CMV-specific CD4(+) effector-memory T helper cells by autologous PDC and induction of IFN-I production in PDC are dependent on appropriate stimulation. Type B CpG oligonucleotide (CpG-B)-stimulated PDC efficiently process and present CMV antigen and are thus capable of stimulating CMV-specific effector-memory T helper cells. CpG-A-stimulated PDC produce large amounts of IFN-I and express programmed death receptor-1 ligand 1. CpG-A plus CpG-B-co-stimulated PDC behave like CpG-B-stimulated PDC, suggesting that antigen processing and presentation in PDC is dependent on stimulation that concurrently inhibits IFN-I production. In vivo targeting of antigens to PDC via CD303 combined with appropriate PDC stimulation may allow induction of specific T cell activation.     

Kinetics and expression patterns of chemokine receptors in human CD4+ T lymphocytes primed by myeloid or plasmacytoid dendritic cells

We investigated the kinetics of expression of 12 chemoattractant receptors as a function of cell division following priming of human naive CD4+ T cells by different populations of dendritic cells (DC) and under conditions favoring Th1 or Th2 differentiation. Two chemokine receptors, CXCR3 and CXCR5, were rapidly up-regulated following T cell activation by either monocyte-derived DC, myeloid DC (mDC) or plasmacytoid DC (pDC). While CXCR5 expression was transient, expression of CXCR3 at advanced cell divisions was dependent on differentiation, being expressed at high levels on Th1 cells. Several other receptors (CCR2, CCR3, CCR4, CCR5, CXCR6 and CRTh2) were acquired progressively as a function of cell division and in a fashion that was influenced by polarizing cytokines. The Th2-associated chemoattractant receptors CRTh2 and CCR3 were up-regulated with slower kinetics compared to the Th1-associated receptors CXCR3 and CXCR6, consistent with a different kinetics and efficiency of polarization. Moreover, CCR4 and CXCR6 were preferentially induced in T cells activated by mDC and pDC, respectively. Finally, CXCR5 and CCR7 were also rapidly and transiently up-regulated in memory T cells following TCR stimulation. These results indicate a complex chemokine receptor regulation dependent on both T cell activation and differentiation state. In addition, they reveal the existence of DC-specific cues for the regulation of T cell migratory capacity.     

HIV-infected cells are major inducers of plasmacytoid dendritic cell interferon production, maturation, and migration

Plasmacytoid dendritic cells (PDC), natural type-1 interferon (IFN) producing cells, could play a role in the innate anti-HIV immune response. Previous reports indicated that PDC IFN production is induced by HIV. Our results show a more robust IFN induction when purified PDC (>95%) were exposed to HIV-infected cells. This effect was not observed with non-viable cells, DNA, and RNA extracted from infected cells, and viral proteins. The response was blocked by anti-CD4 and neutralizing anti-gp120 antibodies as well as soluble CD4. IFN induction by HIV-infected cells was also prevented by low-dose chloroquine, which inhibits endosomal acidification. PDC IFN release resulted in reduced HIV production by infected CD4+ cells, supporting an anti-HIV activity of PDC. Stimulated CD4+ cells induced PDC activation and maturation; markers for PDC migration (CCR7) were enhanced by HIV-infected CD4+ cells only. This latter finding could explain the decline in circulating PDC in HIV-infected individuals.     

Preferential depletion of CD2(low) plasmacytoid dendritic cells in HIV-infected subjects

Plasmacytoid dendritic cells (pDCs) are decreased in number and are functionally impaired in HIV act reasons for pDCs depletion are still unknown. It was recently reported that pDCs can be divided into two functionally distinct populations based on their CD2 expression level. To determine how the CD2(high) and CD2(low) populations are affected by HIV infection, we analyzed their frequencies in the peripheral blood of HIV-infected subjects and healthy controls. We found that the CD2(low) pDC subset was preferentially depleted in infected individuals. The frequency of CD2(low) pDCs correlated with the CD4(+) T-cell count but not with the plasma viral load. This finding furthers our understanding of the causes and consequences of pDC depletion during HIV infection.     

浆细胞样树突状细胞在感染性、免疫性疾病中的研究进展

浆细胞样树突状细胞（PDC）来源于淋巴系造血干／祖细胞,能选择性的诱导免疫应答,并在一定条件下诱导免疫耐受,在抗病毒感染中起重要作用,并可能参与了某些自身免疫性疾病的发生。PDC产生的Ⅰ型IFN（IFN-α／β）在抗病毒过程中很关键,而一定条件下PDC诱导的免疫耐受与调节性T细胞的产生相关。自身免疫性疾病中持续高水平的Ⅰ型IFN提示PDC可能参与此类疾病的发病机制。     

Activation of human NK cells by plasmacytoid dendritic cells and its modulation by CD4+ T helper cells and CD4+ CD25hi T regulatory cells

Plasmacytoid dendritic cells (pDC) represent a specialized cell population that produce type I interferon (IFN) in response to virus. Although type I IFN is a natural killer (NK) cell modulator, a direct role for pDC in coordinating NK cell functions has not yet been elucidated in detail, especially in humans. Here we report that human pDC, following engagement of Toll-like receptor (TLR) 9, not only activate autologous NK cells, as indicated by the induction of CD69 expression, but also enhance their effector functions, especially cytotoxicity. Moreover, they can induce selective proliferation of CD56bright CD16- NK cells. This activity can be strongly augmented by the addition of autologous CD4+ CD25- T helper cells in an IL-2-dependent fashion. Strikingly, CD4+ CD25hi T regulatory (Treg) cells completely abrogate this IL-2-dependent proliferation of NK cells, while they are not able to influence NK cell activation or proliferation solely induced by pDC. Our data show that TLR9-engaged pDC represent a critical stimulus for human NK cells and that CD4+ Th cells and CD4+ CD25hi Treg cells play an important role in modulating human NK cell responses.     

Expression of CD33-related siglecs on human mononuclear phagocytes, monocyte-derived dendritic cells and plasmacytoid dendritic cells

Siglecs are sialic acid binding Ig-like lectins mostly expressed in the haemopoietic and immune systems. Amongst the 11 human siglecs, there are eight proteins highly related to CD33 which have biochemical features of inhibitory receptors, containing two conserved tyrosine-based inhibitory motifs. Five of these (CD33/siglec-3, -5, -7, -9 and -10) are expressed on circulating monocytes. Here we show that monocytes cultured to differentiate into macrophages using either GM-CSF or M-CSF retained expression of these siglecs and their levels were unaffected following stimulation with LPS. In comparison, monocyte-derived dendritic cells down-modulated siglec-7 and -9 following maturation with LPS. Plasmacytoid dendritic cells in human blood expressed siglec-5 only. On monocytes, siglec-5 was shown to mediate rapid uptake of anti-siglec-5 (Fab)2 fragments into early endosomes. This suggests, in addition to inhibitory signalling, a potential role in endocytosis for siglec-5 and the other CD33-related siglecs. Our results show that siglecs are differentially expressed on mononuclear phagocytes and dendritic cells and that some can be modulated by stimuli that promote maturation and differentiation.     

Leukemic plasmacytoid dendritic cells share phenotypic and functional features with their normal counterparts

This work aims to further characterize the newly described leukemic plasmacytoid dendritic cells (LPDC), for which we had previously demonstrated their normal, PDC-like ability to produce IFN-alpha. In addition, LPDC also express the specific antigens BDCA-2 and BDCA-4. Importantly, they become fully competent antigen-presenting cells (APC) after a short maturation induced by IL-3 + CD40L or virus, exhibiting a characteristic APC phenotype (high expression of CD83 and of the costimulatory molecules CD40, CD80, CD86). Whereas IL-3 + CD40L-activated LPDC prime naive CD4(+) T cells towards a Th2 pathway (IL-4-secreting T cells), virus-activated LPDC drive a Th1 profile (IFN-gamma-secreting T cells). Moreover, we show in one case that LPDC are able to capture, process and present exogenous antigens, leading to the activation of both CD4(+) and CD8(+) T cell clones in an antigen-specific manner. This study further characterizes the phenotype and immunological functions of LPDC.     

PACSIN1 regulates the TLR7/9-mediated type I interferon response in plasmacytoid dendritic cells

Plasmacytoid dendritic cells (pDCs) are the professional interferon (IFN)-producing cells of the immune system. pDCs specifically express Toll-like receptor (TLR)7 and TLR9 molecules and produce massive amounts of type I IFN by sensing microbial nucleic acids via TLR7 and TLR9. Here we report that protein kinase C and casein kinase substrate in neurons (PACSIN) 1, is specifically expressed in human and mouse pDCs. Knockdown of PACSIN1 by short hairpin RNA (shRNA) in a human pDC cell line significantly inhibited the type I IFN response of the pDCs to TLR9 ligand. PACSIN1-deficient mice exhibited normal levels of conventional DCs and pDCs, demonstrating that development of pDCs was intact although PACSIN1-deficient pDCs showed reduced levels of IFN-α production in response to both cytosine guanine dinucleotide (CpG)-oligonucleotide (ODN) and virus. In contrast, the production of proinflammatory cytokines in response to those ligands was not affected in PACSIN1-deficient pDCs, suggesting that PACSIN1 represents a pDC-specific adaptor molecule that plays a specific role in the type I IFN signaling cascade.     

CD4+CXCR3+ T cells and plasmacytoid dendritic cells drive accelerated atherosclerosis associated with systemic lupus erythematosus

Cardiovascular disease due to accelerated atherosclerosis is the leading cause of death in patients with systemic lupus erythematosus (SLE). Noteworthy, accelerated atherosclerosis in SLE patients appears to be independant of classical Framingham risk factors. This suggests that aggravated atherosclerosis in SLE patients may be a result of increased inflammation and altered immune responses. However, the mechanisms that mediate the acceleration of atherosclerosis in SLE remain elusive. Based on experimental data which includes both humans (SLE patients and control subjects) and rodents (ApoE−/− mice), we herein propose a multi-step model in which the immune dysfunction associated with SLE (i.e. high level of IFN-α production by TLR 9-stimulated pDCs) is associated with, first, an increased frequency of circulating pro inflammatory CD4+CXCR3+ T cells; second, an increased production of CXCR3 ligands by endothelial cells; third, an increased recruitment of pro-inflammatory CD4+CXCR3+ T cells into the arterial wall, and fourth, the development of atherosclerosis. In showing how SLE may promote accelerated atherosclerosis, our model also points to hypotheses for potential interventions, such as pDCs-targeted therapy, that might be studied in the future.     

CpG ODN enhance antigen-specific NKT cell activation via plasmacytoid dendritic cells

Human Valpha24+ Vbeta11+ natural killer T cells (NKT cells) are "natural memory" T cells that detect glycolipid antigens such as alpha-galactosylceramide (alpha-GalCer) presented on CD1d. In the present study we found that highly purified Valpha24+ NKT cells lack TLR9 mRNA, and thus are not sensitive towards stimulation with CpG oligodeoxynucleotides (ODN). Within PBMC, however, CpG ODN synergistically activated NKT cells stimulated with their cognate antigen alpha-GalCer. Depletion of plasmacytoid dendritic cells (PDC) or myeloid dendritic cells (MDC) revealed that both DC subsets were necessary for the synergistic activation of NKT cells by alpha-GalCer and CpG ODN. While PDC were responsible for the stimulation of NKT cells with CpG ODN, MDC but not PDC presented alpha-GalCer via CD1d. Partial activation of NKT cells was mediated by PDC-derived IFN-alpha, whereas full activation of NKT cells as indicated by IFN-gamma production required cell-to-cell contact of PDC and NKT cells in addition to IFN-alpha; OX40 was involved in this interaction. We conclude that CpG-activated PDC enhance alpha-GalCer-specific NKT cell activation, and bias activated NKT cells towards a Th1 phenotype. Our results lead to a novel concept of PDC function: to regulate effector activity of antigen-stimulated T cells in a cell contact-dependent manner without the need of simultaneous presentation of the cognate T cell antigen.     

CD4+CD56+ hematodermic neoplasms bear a plasmacytoid dendritic cell phenotype

CD4+CD56+ hematodermic neoplasms (HNs) with initial presentation in the skin are characterized by highly aggressive behavior and poor prognosis. Recent studies indicate that malignant cells, which are devoid of common T-, B-, NK-, and myeloid lineage markers, may be of plasmacytoid dendritic cell (pDC) origin. We undertook a study to assess the expression of several pDC-associated molecules on a series of 5 CD4+CD56+ HN cases. CD123 was expressed in all 5 cases, with some heterogeneity in individual cases. All but one case revealed fine membranous BDCA-2 staining of the dermal infiltrate. pDC-like phenotype of the malignant infiltrating cells was confirmed by costaining of BDCA-2+ cells with CD123 and CD4. MxA protein, representing the surrogate marker for lesional type I interferon activity, was expressed in 4 of 5 evaluated cases. Our findings further substantiate the putative pDC origin of CD4+CD56+ HNs.     

IL-2/IL-2R对单核来源树突状细胞的体外分化发育和功能的作用

目的 分别比较抗人CD40单抗和肿瘤坏死因子(tumor necrosis factor,TNF)对单核来源树突状细胞(monocyte-derived dendritic cell,Mo-DC)体外诱导表达CD25(IL-2Rα)的作用,并进一步探讨了IL-2在Mo-DC分化成熟和功能介导中的生物学效应.方法 采用联合粒细胞-巨噬细胞集落刺激因子(granulocyte-macrophage colony stimulating factor,GM-CSF)和IL-4体外诱导Mo-DC的培养方法 ,分别加入CD40单抗和TNF-α诱导Mo-DC成熟;免疫荧光标记分析Mo-DC表型;ELISA法测定γ干扰素(interferon-r,IFN-γ)的分泌水平;3H-TdR掺入法分析Mo-DC对自体T细胞的促增殖效应.结果 (1)CD40信号能有效诱导成熟Mo-DC上调表达CD25(IL-2Rα),而TNF-α则无此作用;(2)Mo-DC在CD40激发后,加入不同剂量的IL-2继续培养可促进Mo-DC对IFN-γ的分泌,同时3H-TdR掺入法的结果也显示,在Mo-DC培养时加入IL-2可以增强Mo-DC对T细胞的促增殖作用.结论 CD40激发的Mo-DC上调表达CD25(IL-2Rα),外源性IL-2能促进Mo-DC分泌IFN-γ和介导T细胞的促增殖效应.     

Vaccination with plasmacytoid dendritic cells induces protection against infection with Leishmania major in mice

DC-based vaccination against Leishmania major induces a parasite-specific Th1 response and long-lasting protective immunity in susceptible mice. Since distinct DC subsets have been proposed to direct the predominant development of either Th1 or Th2 cells, we analyzed the capability of plasmacytoid DC (pDC) to induce protection and elicit a Th1 response against L. major. Pulsing with L. major lysate induced the activation and maturation of semi-mature murine pDC that had been isolated from the spleen, as indicated by up-regulation of the co-stimulatory molecules CD86 and CD80, but did not enhance the level of IFN-alpha secretion by pDC. Vaccination of susceptible mice with L. major lysate-pulsed pDC induced highly effective T cell-mediated immunity against subsequent infection with L. major parasites. Surprisingly, the protection was not accompanied by a polarized Th1 cytokine profile. Co-activation of pDC with CpG-containing oligodeoxynucleotides, which has been shown to be critical for activating the protective potential of myeloid DC, was not required for the protective effect of L. major antigen-pulsed pDC. These findings demonstrate that antigen-loaded pDC are able to induce T cell-mediated protection against a parasite disease and that experimental leishmaniasis is a suitable model to elucidate the mechanisms underlying DC-based vaccination against infections.